2.4 Characterization
of Total MAME by 1-D
and 2-D TLC
1. Silica gel G60 precoated plate, 20 cm  20 cm, 0.25 mm.
2. TLC tank with saturation paper.
3. HPLC-grade solvents: Dichloromethane, petroleum ether, and
diethyl ether (see Note 2).
4. 0.1% rhodamine B in ethanol diluted to 0.01% in 0.25 M
NaH 2 PO 4 solution.
5. 10% molybdophosphoric acid in ethanol.
6. 0.01% primulin in H 2 O/acetone 1:4 v/v.
7. UV light lamp CAMAG for TLC plates.
8. Heater for charring the plates.
9. Pasteur pipettes or glass syringe.
2.5 Characterization
of FAME and MAME
by GC-MS
1. Gas-chromatograph (Trace Thermo Fisher) fitted with a
TG1MS fused-silica capillary column (30 m  0.25 mm) and
connected to an ISQ™ single-quadrupole mass spectrometer
with electron energy of 70 eV. The injection temperature is
fixed at 250
C for performing pyrolysis of MAME.
2. Automatic injector equipped with 10-μL syringe.
3. 1-mL screw-capped glass vials.
2.6 Characterization
and Quantification of
MAME by HPTLC
1. Normal phase high-performance (HP) TLC silica gel
60, 200 μm, 20 Â 10 cm plates.
2. CAMAG ATS4 Automatic TLC sampler 4 equipped of a 25-μL
syringe and N 2 .
3. CAMAG ADC2 Automatic Developing Chamber.
4. CAMAG TLC Scanner 3 and winCATs software.
5. CAMAG Chromatogram Immersion Device III.
6. Saturation sheets.
7. Pasteur pipettes.
8. 1- mL screw-capped glass vials.
9. HPLC-grade solvents: Dichloromethane, petroleum ether,
diethyl ether, ethanol, and chloroform (see Note 2).
10. 0.01% primulin in water/acetone 1/4 v/v for detection.
11. Visualization: 365 nm lamp or revelation system UV.
2.7 Characterization
and Quantification of
Free Mycolic Acids by
UPLC-MS
1. UPLC-MS/MS system: UPLC separation module coupled to
ESI-QTOF Xevo G2-XS mass spectrometer (Waters, Milford,
MA, USA).
2. Acquity UPLC BEH C18 column (2.1 mm  100 mm,
1.7 μm) (Waters).
3. Ultrasonic cleaner.
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Marie-Antoinette Lane ´ elle et al.
of Total MAME by 1-D
and 2-D TLC
1. Silica gel G60 precoated plate, 20 cm  20 cm, 0.25 mm.
2. TLC tank with saturation paper.
3. HPLC-grade solvents: Dichloromethane, petroleum ether, and
diethyl ether (see Note 2).
4. 0.1% rhodamine B in ethanol diluted to 0.01% in 0.25 M
NaH 2 PO 4 solution.
5. 10% molybdophosphoric acid in ethanol.
6. 0.01% primulin in H 2 O/acetone 1:4 v/v.
7. UV light lamp CAMAG for TLC plates.
8. Heater for charring the plates.
9. Pasteur pipettes or glass syringe.
2.5 Characterization
of FAME and MAME
by GC-MS
1. Gas-chromatograph (Trace Thermo Fisher) fitted with a
TG1MS fused-silica capillary column (30 m  0.25 mm) and
connected to an ISQ™ single-quadrupole mass spectrometer
with electron energy of 70 eV. The injection temperature is
fixed at 250
C for performing pyrolysis of MAME.
2. Automatic injector equipped with 10-μL syringe.
3. 1-mL screw-capped glass vials.
2.6 Characterization
and Quantification of
MAME by HPTLC
1. Normal phase high-performance (HP) TLC silica gel
60, 200 μm, 20 Â 10 cm plates.
2. CAMAG ATS4 Automatic TLC sampler 4 equipped of a 25-μL
syringe and N 2 .
3. CAMAG ADC2 Automatic Developing Chamber.
4. CAMAG TLC Scanner 3 and winCATs software.
5. CAMAG Chromatogram Immersion Device III.
6. Saturation sheets.
7. Pasteur pipettes.
8. 1- mL screw-capped glass vials.
9. HPLC-grade solvents: Dichloromethane, petroleum ether,
diethyl ether, ethanol, and chloroform (see Note 2).
10. 0.01% primulin in water/acetone 1/4 v/v for detection.
11. Visualization: 365 nm lamp or revelation system UV.
2.7 Characterization
and Quantification of
Free Mycolic Acids by
UPLC-MS
1. UPLC-MS/MS system: UPLC separation module coupled to
ESI-QTOF Xevo G2-XS mass spectrometer (Waters, Milford,
MA, USA).
2. Acquity UPLC BEH C18 column (2.1 mm  100 mm,
1.7 μm) (Waters).
3. Ultrasonic cleaner.
118
Marie-Antoinette Lane ´ elle et al.